74 THE FROG 
be collected when available, and a supply kept on hand; it 
furnishes a good object with which to begin the study of the 
tissues. Permanent stained mounts are hardly necessary; if 
they are desired, the material should be treated with corrosive- 
acetic, stained with a nuclear stain such as Delafield’s hzmo- 
toxylin, and counterstained with Orange G or with eosin. 
2. Cartilage. Cartilage taken from frogs hardened in formalin 
in the usual way is seldom in good condition, since the formalin 
does not penetrate quickly enough. Material for this study 
should be prepared by placing the fresh femur, or other parts of 
the skeleton containing hyaline cartilage, in strong formalin (10 
per cent.) ; or the fresh cartilage may be examined in normal salt 
solution (for formula see foot-note, p. 45). Free-hand sections 
should be cut with a razor. Such preparations are likely to be 
more satisfactory than permanent stained mounts. 
3. Blood. Permanent preparations are desirable to show more 
clearly the nuclei of the red blood corpuscles. A rapid method is 
is to smear a thin film of blood on a slide, fix it in a saturated 
solution of corrosive sublimate, and stain in Delafield’s hzemo- 
toxylin, counterstaining with Orange G or with eosin. For details 
of this and other methods see Guyer, Animal Micrology. 
4. Non-striated muscle fiber. After ligaturing the intestine of 
a freshly-killed frog, inflate the bladder with air and place it in the 
fixing fluid (corrosive-acetic or Zenker’s) while still inflated; or 
stretch it out on a piece of cork and submerge in the fixing fluid. 
Stain with Delafield’s hemotoxylin, and counterstain with Orange 
G or with eosin. Cut it into pieces of suitable size for mounting. 
5. Connective tissue. As directed in the text, material may 
be taken from a frog hardened in formalin, and studied without 
staining, but there are some advantages in using permanent 
stained preparations for comparison. The thin septa which 
fasten the skin of the frog to the body consist for the most part 
of white fibrous connective tissue; cut out pieces of this from a 
fresh frog, spread on a slide and fix with corrosive sublimate 
(avoid the use of fixing fluids containing acetic acid, since this 
tends to dissolve the white fibers leaving the yellow elastic fibers 
exposed). Stain with Mallory’s connective tissue stain (see 
Guyer, Animal Micrology). 
